Evaluation Of An In-house Specific Immunoglobulin G (igg) Avidity Elisa For Distinguishing Recent Primary From Long-term Human Cytomegalovirus (hcmv) Infection.

This article describes the standardization and evaluation of an in-house specific IgG avidity ELISA for distinguishing recent primary from long-term human cytomegalovirus (HCMV) infection. The test was standardized with the commercial kit ETI-CYTOK G Plus (Sorin Biomedica, Italy) using 8 M urea in phosphate-buffered saline to dissociate low-avidity antibodies after the antigen-antibody interaction. The performance of the in-house assay was compared to that of the commercial automated VIDAS CMV IgG avidity test (bioM rieux, France). Forty-nine sera, 24 from patients with a recent primary HCMV infection and 25 from patients with a long-term HCMV infection and a sustained persistence of specific IgM antibodies, were tested. Similar results were obtained with the two avidity methods. All 24 sera from patients with recently acquired infection had avidity indices compatible with acute HCMV infection by the VIDAS method, whereas with the in-house method, one serum sample had an equivocal result. In the 25 sera from patients with long-term infection, identical results were obtained with the two methods, with only one serum sample having an incompatible value. These findings suggest that our in-house avidity test could be a potentially useful tool for the immunodiagnosis of HCMV infection.45323-

Avaliação de um teste de avidez imunoenzimático para o citomegalovírus humano (ELISA-HCMV) para distinguir a infecção primária recente da infecção de longa duração

This article describes the standardization and evaluation of an in-house specific IgG avidity ELISA for distinguishing recent primary from long-term human cytomegalovirus (HCMV) infection. The test was standardized with the commercial kit ETI-CYTOK G Plus (Sorin Biomedica, Italy) using 8 M urea in phosphate-buffered saline to dissociate low-avidity antibodies after the antigen-antibody interaction. The performance of the in-house assay was compared to that of the commercial automated VIDAS CMV IgG avidity test (bioMérieux, France). Forty-nine sera, 24 from patients with a recent primary HCMV infection and 25 from patients with a long-term HCMV infection and a sustained persistence of specific IgM antibodies, were tested. Similar results were obtained with the two avidity methods. All 24 sera from patients with recently acquired infection had avidity indices compatible with acute HCMV infection by the VIDAS method, whereas with the in-house method, one serum sample had an equivocal result. In the 25 sera from patients with long-term infection, identical results were obtained with the two methods, with only one serum sample having an incompatible value. These findings suggest that our in-house avidity test could be a potentially useful tool for the immunodiagnosis of HCMV infection.Este artigo descreve a padronização e avaliação de um teste de avidez imunoenzimático para o citomegalovírus humano (ELISA-HCMV) para distinguir a infecção primária recente da infecção de longa duração. O teste foi padronizado com o kit comercial ETI-CYTOK G Plus (Sorin Biomedica, Itália), utilizando uréia 8 M para a dissociação dos anticorpos de baixa avidez. A performance do teste ELISA-HCMV foi comparada com a do teste de avidez comercial automatizado VIDAS CMV IgG (bioMérieux, França), utilizando 24 soros de pacientes com infecção primária recente e 25 soros de pacientes com infecção de longa duração apresentando persistência de anticorpos específicos IgM. Resultados similares foram obtidos com os dois métodos de avidez. Todos os 24 soros de pacientes com infecção recentemente adquirida apresentaram índices de avidez compatíveis com infecção aguda pelo HCMV utilizando o teste VIDAS CMV IgG, enquanto que um dos soros apresentou resultado duvidoso no teste ELISA-HCMV. Os 25 soros de pacientes com infecção de longa duração apresentaram resultados idênticos com os dois métodos, com apenas um dos soros apresentando um valor não compatível. Estes resultados sugerem que o teste de avidez descrito pode ser potencialmente útil para o imunodiagnóstico da infecção pelo HCMV

Surveillance of active human cytomegalovirus infection in hematopoietic stem cell transplantation (HLA sibling identical donor): search for optimal cutoff value by real-time PCR

<p>Abstract</p> <p>Background</p> <p>Human cytomegalovirus (CMV) infection still causes significant morbidity and mortality after allogeneic hematopoietic stem cell transplantation (HSCT). Therefore, it is extremely important to diagnosis and monitor active CMV infection in HSCT patients, defining the CMV DNA levels of virus replication that warrant intervention with antiviral agents in order to accurately prevent CMV disease and further related complications.</p> <p>Methods</p> <p>During the first 150 days after allogeneic HSTC, thirty patients were monitored weekly for active CMV infection by <it>pp65 </it>antigenemia, nested-PCR and real-time PCR assays. Receiver operating characteristic (ROC) plot analysis was performed to determine a threshold value of the CMV DNA load by real-time PCR.</p> <p>Results</p> <p>Using ROC curves, the optimal cutoff value by real-time PCR was 418.4 copies/10⁴PBL (sensitivity, 71.4%; specificity, 89.7%). Twenty seven (90%) of the 30 analyzed patients had active CMV infection and two (6.7%) developed CMV disease. Eleven (40.7%) of these 27 patients had acute GVHD, 18 (66.7%) had opportunistic infection, 5 (18.5%) had chronic rejection and 11 (40.7%) died - one died of CMV disease associated with GVHD and bacterial infection.</p> <p>Conclusions</p> <p>The low incidence of CMV disease in HSCT recipients in our study attests to the efficacy of CMV surveillance based on clinical routine assay. The quantification of CMV DNA load using real-time PCR appears to be applicable to the clinical practice and an optimal cutoff value for guiding timely preemptive therapy should be clinically validated in future studies.</p

Acute graft versus host disease

Acute graft-versus-host disease (GVHD) occurs after allogeneic hematopoietic stem cell transplant and is a reaction of donor immune cells against host tissues. Activated donor T cells damage host epithelial cells after an inflammatory cascade that begins with the preparative regimen. About 35%–50% of hematopoietic stem cell transplant (HSCT) recipients will develop acute GVHD. The exact risk is dependent on the stem cell source, age of the patient, conditioning, and GVHD prophylaxis used. Given the number of transplants performed, we can expect about 5500 patients/year to develop acute GVHD. Patients can have involvement of three organs: skin (rash/dermatitis), liver (hepatitis/jaundice), and gastrointestinal tract (abdominal pain/diarrhea). One or more organs may be involved. GVHD is a clinical diagnosis that may be supported with appropriate biopsies. The reason to pursue a tissue biopsy is to help differentiate from other diagnoses which may mimic GVHD, such as viral infection (hepatitis, colitis) or drug reaction (causing skin rash). Acute GVHD is staged and graded (grade 0-IV) by the number and extent of organ involvement. Patients with grade III/IV acute GVHD tend to have a poor outcome. Generally the patient is treated by optimizing their immunosuppression and adding methylprednisolone. About 50% of patients will have a solid response to methylprednisolone. If patients progress after 3 days or are not improved after 7 days, they will get salvage (second-line) immunosuppressive therapy for which there is currently no standard-of-care. Well-organized clinical trials are imperative to better define second-line therapies for this disease. Additional management issues are attention to wound infections in skin GVHD and fluid/nutrition management in gastrointestinal GVHD. About 50% of patients with acute GVHD will eventually have manifestations of chronic GVHD

Withdrawal of maintenance therapy for cytomegalovirus retinitis in AIDS patients exhibiting immunological response to HAART

BACKGROUND: Before the introduction of highly active antiretroviral therapy (HAART), CMV retinitis was a common complication in patients with advanced HIV disease and the therapy was well established; it consisted of an induction phase to control the infection with ganciclovir, followed by a lifelong maintenance phase to avoid or delay relapses. METHODS: To determine the safety of CMV maintenance therapy withdrawal in patients with immune recovery after HAART, 35 patients with treated CMV retinitis, on maintenance therapy, with CD4+ cell count greater than 100 cells/mm³ for at least three months, but almost all patients presented these values for more than six months and viral load < 30000 copies/mL, were prospectively evaluated for the recurrence of CMV disease. Maintenance therapy was withdrawal at inclusion, and patients were monitored for at least 48 weeks by clinical and ophthalmologic evaluations, and by determination of CMV viremia markers (antigenemia-pp65), CD4+/CD8+ counts and plasma HIV RNA levels. Lymphoproliferative assays were performed on 26/35 patients. RESULTS: From 35 patients included, only one had confirmed reactivation of CMV retinitis, at day 120 of follow-up. No patient returned positive antigenemia tests. No correlation between lymphoproliferative assays and CD4+ counts was observed. CONCLUSION: CMV retinitis maintenance therapy discontinuation is safe for those patients with quantitative immune recovery after HAART

Physicochemical Characterization Of Three Fiber-reinforced Epoxide-based Composites For Dental Applications

Fiber-reinforced composite (FRC) biomedical materials are in contact with living tissues arising biocompatibility questions regarding their chemical composition. The hazards of materials such as Bisphenol A (BPA), phthalate and other monomers and composites present in FRC have been rationalized due to its potential toxicity since its detection in food, blood, and saliva. This study characterized the physicochemical properties and degradation profiles of three different epoxide-based materials intended for restorative dental applications. Characterization was accomplished by several methods including FTIR, Raman, Brunauer-Emmett-Teller (BET) Analysis, X-ray fluorescence spectroscopy, and degradation experiments. Physicochemical characterization revealed that although materials presented similar chemical composition, variations between them were more largely accounted by the different phase distribution than chemical composition. (C) 2016 Elsevier B.V. All rights reserved.69905913VJM Ltd

Berberine slows cell growth in autosomal dominant polycystic kidney disease cells.

Autosomal dominant polycystic kidney disease (ADPKD) is the most common hereditary monogenic disorder characterized by development and enlargement of kidney cysts that lead to loss of renal function. It is caused by mutations in two genes (PKD1 and PKD2) encoding for polycystin-1 and polycystin-2 proteins which regulate different signals including cAMP, mTOR and EGFR pathways. Abnormal activation of these signals following PC1 or PC2 loss of function causes an increased cell proliferation which is a typical hallmark of this disease. Despite the promising findings obtained in animal models with targeted inhibitors able to reduce cystic cell growth, currently, no specific approved therapy for ADPKD is available. Therefore, the research of new more effective molecules could be crucial for the treatment of this severe pathology. In this regard, we have studied the effect of berberine, an isoquinoline quaternary alkaloid, on cell proliferation and apoptosis in human and mouse ADPKD cystic cell lines. Berberine treatment slows cell proliferation of ADPKD cystic cells in a dose-dependent manner and at high doses (100μg/mL) it induces cell death in cystic cells as well as in normal kidney tubule cells. However, at 10μg/mL, berberine reduces cell growth in ADPKD cystic cells only enhancing G0/G1 phase of cell cycle and inhibiting ERK and p70-S6 kinases. Our results indicate that berberine shows a selected antiproliferative activity in cellular models for ADPKD, suggesting that this molecule and similar natural compounds could open new opportunities for the therapy of ADPKD patients

Quality Control Of Poly(methyl Methacrylate) To Medical Purpose By Multiple Headspace Extraction

The quality of polymers to be used for medical purposes is evaluated by the concentration of residual compounds in the polymeric matrix, especially by the amount of residual monomer. Residual components of the polymerization of monomer can cause allergies and biological complications (stomatitis, dermatitis, cheilitis, and irritability), also collateral effects for the patient which are evident from the first five years after implant placement and could remain for thirty years more. In dentures, for example, these components are dissolved by the saliva and are fixed to adjacent tissue causing allergic reactions, including burning. Multiple headspace extraction gas chromatography (MHS-GC) has been found to be an analytical technique particularly suitable for quantification of residual monomer in process samples with complex matrix, including solids. The major advantage of MHS-GC is that there is no need to pre-treat the sample prior to analysis. In this work, the methodology used to determine the residual monomer of Methyl Methacrylate (MMA) in a polymer of Poly(Methyl Methacrylate) (PMMA) is presented. The PMMA was produced in a controlled pilot plant scale laboratory, with rigorous experimental conditions to be used for medical purposes (artificial bone). The method includes the formulation of a calibration curve which was obtained by injecting different masses (0 - 30 mg) of a standard MMA solution in the Headspace sample vials and treated at ten extraction step in the HS-GC system. The results showed that through the present method it is possible to recover 98% of MMA from a solid matrix of PMMA. Copyright © 2013, AIDIC Servizi S.r.l.3216991704Chai, X.S., Hou, Q.X., Schork, F.J., Determination of residual monomer in polymer latex by full evaporation headspace gas chromatography (2004) J. Chromatogr. A., 1040, pp. 163-167Kim, Y., Cho, S., Analysis of trace residual 1, 3-butadiene in poly(acrylonitrile-co- butadiene-costyrene) (2011) J. Ind. Eng. Chem., 17, pp. 394-396Christie, Y.K., Lung, B., Darvell, W., Methyl methacrylate in poly(methyl methacrylate)-validation of direct injection gas chromatography (2004) J. Chromatogr. A., 1061, pp. 93-98Haas, S.S., Brauer, G.M., Dickson, G., A characterization of polymethylmethacrylate boné cement (1975) J. Bone Joint Surg., 57 A (3), pp. 380-391Hasenwinkel, J.M., Lautenschlager, E.P., Wixson, R.L., Gilbert, J.L., Effect of initiation chemistry on the fracture toughness, fatigue strength, and residual monomer content of a novel high-viscosity, twosolution acrylic bone cement (2001) J. Biomed. Mater. Res., 59, pp. 411-421Li, H.L., Zhan, H.Y., Fu, S.Y., Liu, M.R., Chai, X.S., Rapid determination of methanol in black liquors by full evaporation headspace gas chromatography (2007) J. Chromatogr. A., 1175, pp. 33-136Li, H.L., Chai, X.S., Deng, Y.L., Zhan, H.Y., Fu, S.Y., Rapid determination of ethanol in fermentation liquors by full evaporation headspace gas chromatography (2009) J. Chromatogr. A., 1216, pp. 169-172Lung, C.Y.K., Darvell, B.W., Minimization of the inevitable residual monomer in denture base acrylic (2005) Dent. Mater., 21, pp. 1119-1128Meier, U., (2009) Determination of Monomers in Polymers by Multiple Headspace Extraction-GC/MS, , http://shop.perkinelmer.com/Content/applicationnotes/ app_determinationmultiple.pdf, accessed 24.09.2012Mikai, M., Koike, M., Fujii, H., Quantitative analysis of allergenic ingredients in eluate extracted from used base resin (2006) J. Oral Rehabil., 33, pp. 216-220Morais, F.A.I., Mello, B.A., Souza, I.A., Ponzi, E.A.C., Revoredo, G.A., Polymers a base methylmethacrylate, Importance in dentistry (2007) Int. J. Dent., 6 (2), pp. 63-66Zhong, J., Chai, X., Qin, X., Fu, S., A full evaporation headspace gas chromatographic method for determination of monomer conversion in cellulose graft poly-methyl methacrylate (2011) Carbohyd. Polym., 86, pp. 367-37